This project aims to study the structure of the heat shock transcription factor (HSF) from yeast, specifically focussing on the complex formed between HSF and its DNA binding site, the heat shock element (HSE). HSF is part of an important homeostatic system called the heat shock response, which serves to protect the cells from damage caused be elevated temperatures and certain other environmental stresses. In yeast, HSF is an essential protein for both constitutive and heat-inducible transcriptional activation, and its transcriptional activity is regulated by post- translational phosphorylations. Furthermore, HSF probably exists as a trimer, a novel oligomeric state for a DNA binding protein, and might contain a new DNA binding motif. Because the basic mechanism of transcriptional regulation of the heat shock response has been highly conserved throughout all eukaryotes, a thorough comprehension of the structure of the yeast HSF will be applicable to our understanding of the heat shock response in all organisms. One of the long term objectives of this research will be to study X-ray crystallography the structure of the complex formed between HSE and a fragment of HSF containing the DNA binding region. To this end, biophysical, biochemical and molecular biological techniques will be used to analyze the sequence requirements for HSE sites and the domain structure and oligomeric state of HSF. With this information, the appropriate oligonucleotide and protein fragments can be prepared for crystallization trials. concurrently, a mutagenic study of the protein determinants necessary for the molecular recognition of the DNA will be initiated. This study will complement the eventual crystal structure.